Ecology Of Influenza A Viruses Research Articles

Expression of influenza A virus glycan receptor candidates in mallard, chicken, and tufted duck.

Influenza A virus (IAV) pandemics result from interspecies transmission events within the avian reservoir and further into mammals including humans. Receptor incompatibility due to differently expressed glycan structures between species has been suggested to limit zoonotic IAV transmission from the wild bird reservoir as well as between different bird species. Using glycoproteomics, we have studied the repertoires of expressed glycan structures with focus on putative sialic acid-containing glycan receptors for IAV in mallard, chicken and tufted duck; three bird species with different roles in the zoonotic ecology of IAV. The methodology used pinpoints specific glycan structures to specific glycosylation sites of identified glycoproteins and was also used to successfully discriminate α2-3- from α2-6-linked terminal sialic acids by careful analysis of oxonium ions released from glycopeptides in tandem MS/MS (MS2), and MS/MS/MS (MS3). Our analysis clearly demonstrated that all three bird species can produce complex N-glycans including α2-3-linked sialyl Lewis structures, as well as both N- and O- glycans terminated with both α2-3- and α2-6-linked Neu5Ac. We also found the recently identified putative IAV receptor structures, Man-6P N-glycopeptides, in all tissues of the three bird species. Furthermore, we found many similarities in the repertoires of expressed receptors both between the bird species investigated and to previously published data from pigs and humans. Our findings of sialylated glycan structures, previously anticipated to be mammalian specific, in all three bird species may have major implications for our understanding of the role of receptor incompatibility in interspecies transmission of IAV.

Wild bird origin Influenza A subtype diversity and reassortment events during the HPAI H5N1 outbreak in Chile, 2022-2023. Proceeding of The First International Avian Influenza Summit, University of Arkansas- October 16-17, 2023"

Highly pathogenic avian influenza (HPAI) H5N1 viruses continue to pose a significant threat to animal and human health worldwide. Since the onset of the HPAI H5N1 outbreak in South America, our research group has been conducting environmental influenza A virus (IAV) surveillance in different wetlands throughout Chile to detect and characterize circulating IAV strains. Between November 2022 and June 2023, we collected and screened 3275 environmental samples for IAV by RT-qPCR targeting the M gene. Of these, 133 were IAV-positive (4.1\%). Further testing by H5-specific RT-qPCR revealed that 14 (10.5\%) of the positive samples corresponded to HPAI H5 2.3.4.4b, collected during the month of November (2), December (8 positives), January (1 positive), March (1 Positive) and June (1 positive). Samples obtained during spring (November and December) were obtained at the Lluta River estuary in the extreme north of Chile. Samples obtained during Summer (January and March) were obtained in Lluta and at Punta Arenas, located in the extreme South of Chile. The sample obtained in Winter (June) was obtained in Maipo, located in Central Chile. Sequence analysis of 6 fully sequenced H5N1 viruses revealed that all segments were closely related to other H5N1 2.3.4.4b viruses circulating at the time in South America. Interestingly, we also obtained several other subtypes of low pathogenic avian influenza viruses (LPAI) during the sampling period: H13N9, H13N8, H12N5, H8N4, H6N8, H6N5, H5N7, H4N6, H2N8 and H2N2. Phylogenetic analysis of these viruses revealed that locally circulating LPAI H12N5, H6N8, and H5N7 subtype viruses, additionally to 4 more viruses without subtype information, had reassorted with HPAI viruses in at least one of the PA and NP segments. The role that these novel reassortant viruses play in the ecology of IAV in Chile and their pathogenic potential is unknown; however, in light of these reassortment events, further surveillance is warranted to detect potentially novel HPAI H5Nx viruses circulating in wild birds in Chile that could have unknown pathogenic and transmission potential.

Epidemiology and Ecology of Influenza A Viruses among Wildlife in the Arctic.

Arctic regions are ecologically significant for the environmental persistence and geographic dissemination of influenza A viruses (IAVs) by avian hosts and other wildlife species. Data describing the epidemiology and ecology of IAVs among wildlife in the arctic are less frequently published compared to southern temperate regions, where prevalence and subtype diversity are more routinely documented. Following PRISMA guidelines, this systematic review addresses this gap by describing the prevalence, spatiotemporal distribution, and ecological characteristics of IAVs detected among wildlife and the environment in this understudied region of the globe. The literature search was performed in PubMed and Google Scholar using a set of pre-defined search terms to identify publications reporting on IAVs in Arctic regions between 1978 and February 2022. A total of 2125 articles were initially screened, 267 were assessed for eligibility, and 71 articles met inclusion criteria. IAVs have been detected in multiple wildlife species in all Arctic regions, including seabirds, shorebirds, waterfowl, seals, sea lions, whales, and terrestrial mammals, and in the environment. Isolates from wild birds comprise the majority of documented viruses derived from wildlife; however, among all animals and environmental matrices, 26 unique low and highly pathogenic subtypes have been characterized in the scientific literature from Arctic regions. Pooled prevalence across studies indicates 4.23% for wild birds, 3.42% among tested environmental matrices, and seroprevalences of 9.29% and 1.69% among marine and terrestrial mammals, respectively. Surveillance data are geographically biased, with most data from the Alaskan Arctic and many fewer reports from the Russian, Canadian, North Atlantic, and Western European Arctic. We highlight multiple important aspects of wildlife host, pathogen, and environmental ecology of IAVs in Arctic regions, including the role of avian migration and breeding cycles for the global spread of IAVs, evidence of inter-species and inter-continental reassortment at high latitudes, and how climate change-driven ecosystem shifts, including changes in the seasonal availability and distribution of dietary resources, have the potential to alter host–pathogen–environment dynamics in Arctic regions. We conclude by identifying gaps in knowledge and propose priorities for future research.

Maintenance and dissemination of avian-origin influenza A virus within the northern Atlantic Flyway of North America.

Wild waterbirds, the natural reservoirs for avian influenza viruses, undergo migratory movements each year, connecting breeding and wintering grounds within broad corridors known as flyways. In a continental or global view, the study of virus movements within and across flyways is important to understanding virus diversity, evolution, and movement. From 2015 to 2017, we sampled waterfowl from breeding (Maine) and wintering (Maryland) areas within the Atlantic Flyway (AF) along the east coast of North America to investigate the spatio-temporal trends in persistence and spread of influenza A viruses (IAV). We isolated 109 IAVs from 1,821 cloacal / oropharyngeal samples targeting mallards (Anas platyrhynchos) and American black ducks (Anas rubripes), two species having ecological and conservation importance in the flyway that are also host reservoirs of IAV. Isolates with >99% nucleotide similarity at all gene segments were found between eight pairs of birds in the northern site across years, indicating some degree of stability among genome constellations and the possibility of environmental persistence. No movement of whole genome constellations were identified between the two parts of the flyway, however, virus gene flow between the northern and southern study locations was evident. Examination of banding records indicate direct migratory waterfowl movements between the two locations within an annual season, providing a mechanism for the inferred viral gene flow. Bayesian phylogenetic analyses provided evidence for virus dissemination from other North American wild birds to AF dabbling ducks (Anatinae), shorebirds (Charidriformes), and poultry (Galliformes). Evidence was found for virus dissemination from shorebirds to gulls (Laridae), and dabbling ducks to shorebirds and poultry. The findings from this study contribute to the understanding of IAV ecology in waterfowl within the AF.

Exposure of domestic swine to influenza A viruses in Ghana suggests unidirectional, reverse zoonotic transmission at the human-animal interface.

Influenza A viruses (IAVs) have both zoonotic and anthroponotic potential and are of public and veterinary importance. Swine are intermediate hosts and 'mixing vessels' for generating reassortants, progenies of which may harbour pandemic propensity. Swine handlers are at the highest risk of becoming infected with IAVs from swine but there is little information on the ecology of IAVs at the human-animal interface in Africa. We analysed and characterized nasal and throat swabs from swine and farmers respectively, for IAVs using RT-qPCR, from swine farms in the Ashanti region, Ghana. Sera were also analysed for IAVs antibodies and serotyped using ELISA and HI assays. IAV was detected in 1.4% (n=17/1,200) and 2.0% (n=2/99) of swine and farmers samples, respectively. Viral subtypes H3N2 and H1N1pdm09 were found in human samples. All virus-positive swine samples were subtyped as H1N1pdm09 phylogenetically clustering closely with H1N1pdm09 that circulated among humans during the study period. Phenotypic markers that confer sensitivity to Oseltamivir were found. Serological prevalence of IAVs in swine and farmers by ELISA was 3.2% (n=38/1,200) and 18.2% (n=18/99), respectively. Human H1N1pdm09 and H3N2 antibodies were found in both swine and farmers sera. Indigenous swine influenza A viruses and/or antibodies were not detected in swine or farmers samples. Majority (98%, n=147/150) of farmers reported of not wearing surgical mask and few (4%, n=6) reported to wear gloves when working. Most (n=74, 87.7%) farmers reported of working on the farm when experiencing influenza-like illness. Poor husbandry and biosafety practices of farmers could facilitate virus transmission across the human-swine interface. Farmers should be educated on the importance of good farm practices to mitigate influenza transmission at the human-animal interface.

Effects of freshwater crayfish on influenza A virus persistence in water.

Several investigations have recently assessed the ability of some aquatic invertebrates to act as tools for avian influenza A virus (IAV) surveillance as well as their potential role(s) in IAV ecology. Because of this, as well as the high IAV seroprevalence rates noted in select mesocarnivores that commonly inhabit aquatic and semi-aquatic habitats, we evaluated the effects that freshwater crayfish have on IAV in water at three dose levels and monitored for the presence of IAV in crayfish tissues (gill and green gland) and haemolymph at multiple time points. At relatively high, medium and low (approximately 104 , 103 and 102 EID50 /ml, respectively) doses, mesocosms containing crayfish (Orconectes sp.) had less detectable IAV RNA present when final water samples were assayed (9days post-contact [DPC]). In general, containers without crayfish present had nearly three-fold greater quantities of viral RNA at 9DPC. A varying number of RNA positive samples were detected for the three crayfish sample types collected. Gill tissue produced the largest number of positive non-water samples (n=26), with the highest quantities detected from crayfish sampled on 1 and 4DPC (103.5 EID50 equivalent/ml). On a few occasions, gill (n=8) and haemolymph samples (n=1) produced higher quantities of viral RNA than their respective water samples or water samples collected 1-2DPC earlier, but these differences were typically minor. Based upon water samples, statistical models indicated that the interaction of dose and crayfish exposure days explained most of the variation in these data. Future efforts should address if crayfish exposed to IAV-laden water have the capacity to successfully transmit IAVs to mammals and birds which frequently prey upon them.

Ecology of Influenza A Viruses in Wild Birds and Wetlands of Alaska.

Alaska represents a globally important region for the ecology of avian-origin influenza A viruses (IAVs) given the expansive wetlands in this region, which serve as habitat for numerous hosts of IAVs that disperse among four continents during the annual cycle. Extensive sampling of wild birds for IAVs in Alaska since 1991 has greatly extended inference regarding intercontinental viral exchange between North America and East Asia and the importance of Beringian endemic species to IAV ecology within this region. Data on IAVs in aquatic birds inhabiting Alaska have also been useful for helping to establish global patterns of prevalence in wild birds and viral dispersal across the landscape. In this review, we summarize the main findings from investigations of IAVs in wild birds and wetlands of Alaska with the aim of providing readers with an understanding of viral ecology within this region. More specifically, we review viral detections, evidence of IAV exposure, and genetic characterization of isolates derived from wild bird samples collected in Alaska by host taxonomy. Additionally, we provide a short overview of wetland complexes within Alaska that may be important to IAV ecology at the continental scale.

Individual-Level Antibody Dynamics Reveal Potential Drivers of Influenza A Seasonality in Wild Pig Populations

Swine are important in the ecology of influenza A virus (IAV) globally. Understanding the ecological role of wild pigs in IAV ecology has been limited because surveillance in wild pigs is often for antibodies (serosurveillance) rather than IAVs, as in humans and domestic swine. As IAV antibodies can persist long after an infection, serosurveillance data are not necessarily indicative of current infection risk. However, antibody responses to IAV infections cause a predictable antibody response, thus time of infection can be inferred from antibody levels in serological samples, enabling identification of risk factors of infection at estimated times of infection. Recent work demonstrates that these quantitative antibody methods (QAMs) can accurately recover infection dates, even when individual-level variation in antibody curves is moderately high. Also, the methodology can be implemented in a survival analysis (SA) framework to reduce bias from opportunistic sampling. Here we integrated QAMs and SA and applied this novel QAM-SA framework to understand the dynamics of IAV infection risk in wild pigs seasonally and spatially, and identify risk factors. We used national-scale IAV serosurveillance data from 15 US states. We found that infection risk was highest during January-March (54% of 61 estimated peaks), with 24% of estimated peaks occurring from May to July, and some low-level of infection risk occurring year-round. Time-varying IAV infection risk in wild pigs was positively correlated with humidity and IAV infection trends in domestic swine and humans, and did not show wave-like spatial spread of infection among states, nor more similar levels of infection risk among states with more similar meteorological conditions. Effects of host sex on IAV infection risk in wild pigs were generally not significant. Because most of the variation in infection risk was explained by state-level factors or infection risk at long-distances, our results suggested that predicting IAV infection risk in wild pigs is complicated by local ecological factors and potentially long-distance translocation of infection. In addition to revealing factors of IAV infection risk in wild pigs, our framework is broadly applicable for quantifying risk factors of disease transmission using opportunistic serosurveillance sampling, a common methodology in wildlife disease surveillance. Future research on the factors that determine individual-level antibody kinetics will facilitate the design of serosurveillance systems that can extract more accurate estimates of time-varying disease risk from quantitative antibody data.

Emperor geese (Anser canagicus) are exposed to a diversity of influenza A viruses, are infected during the non-breeding period and contribute to intercontinental viral dispersal.

Emperor geese (Anser canagicus) are endemic to coastal areas within Beringia and have previously been found to have antibodies to or to be infected with influenza A viruses (IAVs) in Alaska. In this study, we use virological, serological and tracking data to further elucidate the role of emperor geese in the ecology of IAVs in Beringia during the non-breeding period. Specifically, we assess evidence for: (a) active IAV infection during spring staging, autumn staging and wintering periods; (b) infection with novel Eurasian-origin or interhemispheric reassortant viruses; (c) contemporary movement of geese between East Asia and North America; (d) previous exposure to viruses of 14 haemagglutinin subtypes, including Eurasian lineage highly pathogenic (HP) H5 IAVs; and (e) subtype-specific antibody seroconversion and seroreversion. Emperor geese were found to shed IAVs, including interhemispheric reassortant viruses, throughout the non-breeding period; migrate between Alaska and the Russian Far East prior to and following remigial moult; have antibodies reactive to a diversity of IAVs including, in a few instances, Eurasian lineage HP H5 IAVs; and exhibit relatively broad and stable patterns of population immunity among breeding females. Results of this study suggest that emperor geese may play an important role in the maintenance and dispersal of IAVs within Beringia during the non-breeding period and provide information that may be used to further optimize surveillance activities focused on the early detection of Eurasian-origin IAVs in North America.

Serologic Evidence for Influenza a Virus Exposure in Three Loon Species Breeding in Alaska, USA

Limited information exists about exposure to influenza A viruses (IAVs) in many wild waterbird species, including loons. We analyzed serum samples from breeding adult Pacific (Gavia pacifica), Red-throated (Gavia stellata), and Yellow-billed (Gavia adamsii) loons sampled at three locations along the coast of Alaska, US from 2008 to 2017 to gain a better understanding of the potential role loons play in IAV ecology. We screened loon sera for IAV antibodies using three tests-blocking enzyme-linked immunosorbent assay (bELISA), agar gel immunodiffusion (AGID), and hemagglutination inhibition (HI)-and examined patterns in seroprevalence among species and sampling locations. We found evidence of IAV infection in all loon species and at all breeding locations, although concordance was imperfect among serological tests. Diagnostic tests yielded seroprevalence estimates of 24% (42/172) with bELISA, 8% (5/60) with AGID, and 6% (4/70) with HI. The IAV subtypes to which loon sera reacted using HI were consistent with those detected in waterfowl and gulls at other locations in Alaska, suggesting that loons may be exposed to IAV maintained in sympatric waterbirds. Our study provided evidence that loons inhabiting Alaska were exposed to IAV. However, given imperfect concordance among serologic tests, and relatively low seroprevalence as compared to other avian taxa exposed to IAV in Alaska, they make poor IAV surveillance targets.

Adaptation of Human Influenza Viruses to Swine.

A large diversity of influenza A viruses (IAV) within the H1N1/N2 and H3N2 subtypes circulates in pigs globally, with different lineages predominating in specific regions of the globe. A common characteristic of the ecology of IAV in swine in different regions is the periodic spillover of human seasonal viruses. Such human viruses resulted in sustained transmission in swine in several countries, leading to the establishment of novel IAV lineages in the swine host and contributing to the genetic and antigenic diversity of influenza observed in pigs. In this review we discuss the frequent occurrence of reverse-zoonosis of IAV from humans to pigs that have contributed to the global viral diversity in swine in a continuous manner, describe host-range factors that may be related to the adaptation of these human-origin viruses to pigs, and how these events could affect the swine industry.

Influenza A Prevalence and Subtype Diversity in Migrating Teal Sampled Along the United States Gulf Coast.

Wild birds in the order Anseriformes are important reservoirs for influenza A viruses (IAVs); however, IAV prevalence and subtype diversity may vary by season, even at the same location. To better understand the ecology of IAV during waterfowl migration through the Gulf Coast of the United States (Louisiana and Texas), surveillance of blue-winged (Spatula discors) and American green-winged (Anas carolinensis) teal was conducted. The surveillance was done annually during the spring (live capture; 2012-17) and fall (hunter harvested; 2007-17) at times inferred to coincide with northward and southward movements, respectively, for these waterfowl species. During spring migration, 266 low pathogenicity (LP) IAV positive samples were recovered from 7547 paired cloacal-oropharyngeal (COP) samples (prevalence, 3.5%; annual range, 1.3%-8.4%). During fall migration, 650 LP IAV-positive samples were recovered from 9493 COP samples (prevalence, 6.8%; annual range, 0.4%-23.5%). Overall, 34 and 20 different IAV subtypes were recovered during fall and spring sampling, respectively. Consistent with previous results for fall migrating ducks, H3 and H4 hemagglutinin (HA) subtypes were most common; however, H4 subtype viruses predominated every year. This is in contrast to the predominance of LP H7 and H10 HA subtype viruses during spring. The N6 and N8 neuraminidase subtypes, which were usually associated with H4, were most common during fall; the N6 subtype was not recovered in the spring. These consistent seasonal trends in IAV subtype detection in teal are currently not understood and highlight the need for further research regarding potential drivers of spatiotemporal patterns of infection, such as population immunity.

Identifying Gaps in Wild Waterfowl Influenza A Surveillance in Ohio, United States.

The Mississippi Flyway is of utmost importance in monitoring influenza A viral diversity in the natural reservoir, as it is used by approximately 40% of North American migratory waterfowl. In 2008, influenza A virus (IAV) surveillance was initiated in eight states within the flyway during annual southern migration, to gain better insight into the natural history of influenza A viruses in the natural reservoir. More than 45,000 samples have been collected and tested, resulting in hundreds of diverse influenza A viral isolates, but seasonal sampling may not be the best strategy to gain insight into the natural history of IAV. To investigate the progress of this sampling strategy toward understanding the ecology of IAV in wild waterfowl, data from mallard ducks (Anas platyrhynchos) sampled nearly year-round in Ohio were examined. Overall, 3,645 samples were collected from mallards in Ohio from 2008 to 2016, with IAV being recovered from 13.6% of all samples collected. However, when data from each month are examined individually, it becomes apparent that the aggregated summary may be providing a misleading view of IAV in Ohio mallards. For instance, in August the frequency of viral recovery is 29.8%, with isolates representing at least 47 hemagglutinin/ neuraminidase (HA/NA) combinations. In November, during the height of southern migration, IAV isolation drops to 6.2%, with only 25 HA/NA combinations being represented. Our biased sampling towards convenience and high IAV recovery has created gaps in the data set, which prohibit a full understanding of the IAV ecology in this waterfowl population.

Serologic and Virologic Evidence of Influenza A Viruses in Wild Boars ( Sus scrofa) from Two Different Locations in Italy.

Swine influenza viruses (SIVs) have been repeatedly demonstrated to circulate in wild boar ( Sus scrofa) populations, whereas no evidence of exposure to avian influenza viruses (AIVs) has been described in wild boar. To better understand how different environments may influence the ecology of influenza A viruses (IAVs) in wild suid populations, we examined biologic samples of wild boars from two study areas represented by an upland (UL) and a wetland (WL) in northern and central Italy, respectively. Serum samples were collected from 388 wild boars sampled in the UL, whereas both a serum sample and a nasal swab were obtained from each of 35 wild boars sampled in the WL. Twenty of 388 (5.2%) sera from the UL were positive by enzyme-linked immunosorbent assay for the presence of antibodies against influenza A nucleoprotein and some of these samples showed antibodies by hemagglutination inhibition to SIVs of H1N1 (1/20), H1N2 (10/20), and H3N2 (1/20) antigenic subtypes. No IAV-seropositive wild boar was detected in the WL, although one of 35 animals was found to be IAV-positive by both a reverse transcriptase PCR and a real-time reverse transcriptase PCR. We hypothesize an SIV exposure for IAV-seropositive wild boars occupying the UL, whereas a possible AIV spillover from aquatic bird species-natural reservoirs of IAVs-to wild boars in the WL cannot be ruled out. Further research is needed to better understand the role played by wild boars in IAV ecology in Mediterranean habitats.

Prospective surveillance for influenza A virus in Chinese swine farms

Pork production in China is rapidly increasing and swine production operations are expanding in size and number. However, the biosecurity measures necessary to prevent swine disease transmission, particularly influenza A viruses (IAV) that can be zoonotic, are often inadequate. Despite this risk, few studies have attempted to comprehensively study IAV ecology in swine production settings. Here, we present environmental and animal sampling data collected in the first year of an ongoing five-year prospective epidemiological study to assess IAV ecology as it relates to swine workers, their pigs, and the farm environment. From March 2015 to February 2016, we collected 396 each of environmental swab, water, bioaerosol, and fecal/slurry samples, as well as 3300 pig oral secretion samples from six farms in China. The specimens were tested with molecular assays for IAV. Of these, 46 (11.6%) environmental swab, 235 (7.1%) pig oral secretion, 23 (5.8%) water, 20 (5.1%) bioaerosol, and 19 (4.8%) fecal/slurry specimens were positive for influenza A by qRT-PCR. Risk factors for IAV detection among collected samples were identified using bivariate logistic regression. Overall, these first year data suggest that IAV is quite ubiquitous in the swine production environment and demonstrate an association between the different types of environmental sampling used. Given the mounting evidence that some of these viruses freely move between pigs and swine workers, and that mixing of these viruses can yield progeny viruses with pandemic potential, it seems imperative that routine surveillance for novel IAVs be conducted in commercial swine farms.

First detection of influenza A virus genes from wild raccoons in Japan.

Serological surveys have shown that wild raccoons are exposed to influenza A viruses (IAVs); however, no genetic evidence for this IAV infection has been found. In the present study, we first detected IAV genes in wild raccoons captured during periods other than the wintering season of migratory waterfowl and epidemic season of influenza in Japan. Viral matrix (M) and nucleoprotein (NP) genes were detected by a conventional reverse transcription-polymerase chain reaction assay from three suckling siblings and one juvenile without any noticeable clinical signs, although the NP gene could not be detected from one sibling. The sequences of M gene fragments detected from the rectal swabs of three suckling siblings were comparable with each other but different from those detected from the nasal swab of the juvenile raccoon caught from a different site. The sequences of NP gene fragments detected from two suckling siblings were also comparable. These genetic evidences suggest that IAV is maintained among raccoon populations in the northern part of Japan. Further genetic and virological investigation of IAV infection in wild raccoons is needed to better understand the IAV ecology in the field.

Challenges of influenza A viruses in humans and animals and current animal vaccines as an effective control measure.

Influenza A viruses (IAVs) are genetically diverse and variable pathogens that share various hosts including human, swine, and domestic poultry. Interspecies and intercontinental viral spreads make the ecology of IAV more complex. Beside endemic IAV infections, human has been exposed to pandemic and zoonotic threats from avian and swine influenza viruses. Animal health also has been threatened by high pathogenic avian influenza viruses (in domestic poultry) and reverse zoonosis (in swine). Considering its dynamic interplay between species, prevention and control against IAV should be conducted effectively in both humans and animal sectors. Vaccination is one of the most efficient tools against IAV. Numerous vaccines against animal IAVs have been developed by a variety of vaccine technologies and some of them are currently commercially available. We summarize several challenges in control of IAVs faced by human and animals and discuss IAV vaccines for animal use with those application in susceptible populations.

GENETIC CHARACTERIZATION OF H13 AND H16 INFLUENZA A VIRUSES IN GULLS (LARUS SPP.) WITH CLINICALLY SEVERE DISEASE AND CONCURRENT CIRCOVIRUS INFECTION.

Influenza A viruses (IAVs) of the subtypes H13 and H16 are primarily found in gulls ( Larus spp., order Charadriiformes). Although the gull-adapted subtypes replicate efficiently during infection, gulls usually remain apparently healthy during infection. Avian influenza virus isolates are generally separated into two distinct populations, North American and Eurasian, because of the limited gene flow between the continents. Reassortment between these lineages does occur occasionally; however, direct intercontinental transmission of all eight gene segments is rare. Extensive research has been done to understand the ecology of IAV subtypes that naturally circulate in ducks (order Anseriformes), but the ecology of H13 and H16 IAVs in gulls remains far less studied. In Finland, gulls were screened for IAVs for passive (dead and diseased gulls) and active (clinically healthy gulls) surveillance purposes during the years 2005-10. During that period, 11 H13, two H16 viruses, and one H3N8 IAV were detected. We sequenced partial and full-length hemagglutinin genes of these gull-origin IAVs for phylogenetic assessments. All but one of the H13 genes clustered together with northern European and northeastern Asian viruses, whereas one virus clustered with North American viruses. Interestingly, a high rate (10/14) of these low-pathogenic IAVs was detected in dead or diseased gulls. The atypical clinical status of the IAV-positive gulls and previous observations of circovirus-like inclusion bodies in diseased gulls during autopsies, led us to screen for concurrent circovirus infections in our samples. The DNA of circovirus, an immunosuppressive pathogen of both birds and mammals, was detected in 54% (7/13) of the tested IAV-positive gulls, whereas only 25% (14/56) of our panel of IAV-negative gulls tested positive by circovirus PCR.

Evidence of Intercontinental Spread and Uncommon Variants of Low-Pathogenicity Avian Influenza Viruses in Ducks Overwintering in Guatemala.

Over a hundred species of aquatic birds overwinter in Central America's wetlands, providing opportunities for the transmission of influenza A viruses (IAVs). To date, limited IAV surveillance in Central America hinders our understanding of the evolution and ecology of IAVs in migratory hosts within the Western Hemisphere. To address this gap, we sequenced the genomes of 68 virus isolates obtained from ducks overwintering along Guatemala's Pacific Coast during 2010 to 2013. High genetic diversity was observed, including 9 hemagglutinin (HA) subtypes, 7 neuraminidase (NA) subtypes, and multiple avian IAV lineages that have been detected at low levels (<1%) in North America. An unusually large number of viruses with the rare H14 subtype were identified (n = 14) over two consecutive seasons, the highest number of H14 viruses ever reported in a single location, providing evidence for a possible H14 source population located outside routinely sampled regions of North America. Viruses from Guatemala were positioned within minor clades divergent from the main North American lineage on phylogenies inferred for the H3, H4, N2, N8, PA, NP, and NS segments. A time-scaled phylogeny indicates that a Eurasian virus PA segment introduced into the Americas in the early 2000s disseminated to Guatemala during ~2007.1 to 2010.4 (95% highest posterior density [HPD]). Overall, the diversity detected in Guatemala in overwintering ducks highlights the potential role of Central America in the evolution of diverse IAV lineages in the Americas, including divergent variants rarely detected in the United States, and the importance of increasing IAV surveillance throughout Central America. IMPORTANCE Recent outbreaks of highly pathogenic H7N3, H5Nx, and H7N8 avian influenza viruses in North America were introduced by migratory birds, underscoring the importance of understanding how wild birds contribute to the dissemination and evolution of IAVs in nature. At least four of the main IAV duck host species in North America migrate through or overwinter within a narrow strip of Central America, providing opportunities for diverse IAV lineages to mix and exchange gene segments. By obtaining whole-genome sequences of 68 IAV isolates collected from migratory waterfowl in Guatemala (2010 to 2013), the largest data set available from Central America to date, we detected extensive viral diversity, including gene variants rarely found in North America and gene segments of Eurasian origin. Our findings highlight the need for increased IAV surveillance across the geographical span of bird migration flyways, including Neotropical regions that have been vastly undersampled to date.

Influenza A virus vaccines for swine

Economic losses due to influenza A virus (IAV) infections are substantial and a global problem, ranking among the top three major health challenges in the swine industry. Currently, H1 and H3 subtypes circulate in pigs globally associated with different combinations of N1 and N2 subtypes; however, the origin, gene constellation, and antigenic makeup of IAV vary greatly on different continents. Vaccination is one means of mitigating the effects of IAV disease, and vaccines are most effective if the strains included closely match the currently circulating strains in pigs. Genetic analyses provide panoramic views of the virus landscape at the sequence level and, thus, can aid in the selection of well-matched swine IAV vaccine strains, but is not sufficient alone. Additionally, a major challenge in selecting appropriate swine IAV vaccine strains is the co-circulation of multiple lineages of viruses in the same region, requiring multivalent or broadly cross-reacting antigens. Due to this complex IAV ecology in swine, new vaccination strategies and vaccine platforms are needed. The hemagglutinin (HA) viral protein is the major target of neutralizing antibodies, which are widely considered to be correlated with protection. Virus variants that are not recognized by previously elicited antibodies can render traditional vaccines that primarily elicit humoral responses ineffective, and therefore result in the need for vaccine strain reformulation and re-vaccination. In the future, new vaccine platforms may be on the market that will provide alternative options to those currently available. Nonetheless, a collaborative approach is needed to improve IAV vaccine strain selection for use in swine.